Iron-sulfur clusters of biotin synthase in vivo: A Mössbauer study

Biotin synthase, the enzyme that catalyzes the last step of the biosynthesis of biotin, contains only [2Fe-2S]²⁺ clusters when isolated under aerobic conditions. Previous results showed that reconstitution with an excess of FeCl3 and Na₂S under reducing and anaerobic conditions leads to either [4Fe4S]²⁺, [4Fe-4S]⁺, or a mixture of [4Fe-4S]²⁺ and [2Fe-2S]²⁺ clusters. To determine whether any of these possibilities or other different cluster configuration could correspond to the physiological in vivo state, we have used ⁵⁷Fe Mössbauer spectroscopy to investigate the clusters of biotin synthase in whole cells. The results show that, in aerobically grown cells, biotin synthase contains a mixture of [4Fe-4S]²⁺ and [2Fe-2S]²⁺ clusters. A mixed 14Fe-4S]²⁺:[2Fe-2S]²⁺ cluster form has already been observed under certain in vitro conditions, and it has been proposed that both clusters might each play a significant role in the mechanism of biotin synthase. Their presence in vivo is now another argument in favor of this mixed cluster form.